title: Lack of MD-2 expression in human corneal epithelial cells is an underlying mechanism of lipopolysaccharide (LPS) unresponsiveness.
abstract: In the present study we tested the responsiveness of human corneal epithelial cells (HCECs) and corneal fibroblasts to lipopolysaccharide (LPS), a Toll-like receptor (TLR) 4 ligand. Purified Pseudomonas aeruginosa LPS was used to stimulate telomerase-immortalized HCECs (HUCL) and stromal fibroblast (THK) cell lines. Exposure of cells to LPS induced a time-dependent activation of NF-kappaB in THK but not in HUCL cells, as assessed by an increase in IkappaB-alpha phosphorylation and degradation. Concomitant with NF-kappaB activation, LPS-treated THK cells, but not HUCL cells, produced a significantly larger number of cytokines than control untreated cells. A cell surface biotinylation assay revealed that HUCL cells express TLR4 intracellularly, whereas TLR5 is expressed on the cell surface. Furthermore, reverse transcriptase-PCR analysis revealed that HUCL and primary HCECs, in contrast to THK cells, do not express myeloid differentiation (MD)-2. Thus, our results demonstrate that the LPS unresponsiveness of HCECs might be due to deficient expression of MD-2, an essential component for LPS-TLR4 signaling.
authors.affiliation: Department of Ophthalmology, Kresge Eye Institute, Wayne State University School of Medicine, Detroit, MI 48201, USA.
authors.forename: Jing
authors.initials: J
authors.lastname: Zhang
authors.forename: Ashok
authors.initials: A
authors.lastname: Kumar
authors.forename: Michelle
authors.initials: M
authors.lastname: Wheater
authors.forename: Fu-Shin X
authors.initials: FS
authors.lastname: Yu
mesh_terms: D015153:Blotting, Western; D002478:Cells, Cultured; D003315:Cornea / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D015536:Down-Regulation / Q000187:drug effects; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D008070:Lipopolysaccharides / Q000276:immunology / Q000494:pharmacology*; D051218:Lymphocyte Antigen 96 / Q000378:metabolism*; D011550:Pseudomonas aeruginosa / Q000276:immunology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction
chemical_list: D008070:Lipopolysaccharides; D051218:Lymphocyte Antigen 96